LOCATION AND RELEASE OF HYALURONIDASE FROM PORCINE SPERM
Abstract
Hyaluronidase release from boar sperm was examined during the ionophore-induced acrosome reaction. The effects of seminal plasma coating proteins and composition of the medium on hyaluronidase release during cold shock were also examined. Hyaluronidase was not released from boar sperm incubated with 1.0 (mu)M A23187 until vesiculation of the plasma and acrosomal membrane occurred in some sperm. Hyaluronidase release was positively correlated with aryl sulfatase release. In the presence of 1.0 (mu)M ionophore A23187, caffeine accelerated both the vesiculation of the sperm membranes and the release of hyaluronidase. Caffeine had no effect alone. Heated rabbit serum also increased the release of hyaluronidase during incubation for four hours. Vesiculation of the membranes occurred in a few sperm exposed to heated serum. Ejaculated sperm did not readily release hyaluronidase in response to hypoosmolarity or EDTA and acid pH. Ejaculated sperm from intact and vesiculectomized boars released significant amounts of hyaluronidase with cold shock, but sperm from both types of boars were equally susceptible to cold shock. Epididymal sperm, which lacked seminal plasma coating proteins, released less hyaluronidase during cold shock than ejaculated sperm. These results were consistent with the previously observed reduced susceptibility of epididymal sperm to cold shock. Preincubation in seminal plasma did not affect release of hyaluronidase from ejaculated sperm of vesiculectomized boars during cold shock. Preincubation in seminal plasma from vasectomized or intact boars decreased the release of hyaluronidase rom epididymal sperm during cold shock. This appears contradictory to the observation that ejaculated sperm were more sensitive to cold shock than epididymal sperm. Evidently, there is a difference between ejaculated sperm and epididymal sperm plus seminal plasma that is not yet understood. BSA, PVP, EDTA, Tris, epinephrine, thyroxine, and spermine did not affect the release of boar sperm hyaluronidase. In all three treatments which caused release of hyaluronidase (1.0 (mu)M A23187, 50% heated serum, and cold shock) vesiculation or damage to sperm membrane was detectable by electron microscopy. Coating of ejaculated sperm with seminal plasma proteins did not affect the release of hyaluronidase during cold shock or during cold shock in the presence of EDTA. These observations strongly suggest hyaluronidase is not located on or in the plasma membrane of porcine sperm. Porcine sperm hyaluronidase existed in at least two forms with estimated molecular weights of 60,000 and 125,000. Approximately one-fourth of the extracted hyaluronidase activity was present in the higher molecular weight form. Mercaptoethanol (1mM) reduced the quantity of the higher molecular weight form to a negligible amount.
Degree
Ph.D.
Subject Area
Livestock
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